ATP:citrate lyase of Rhodotorula gracilis: purification and properties.
ATP:citrate lyase was purified from the oleaginous yeast Rhodotorula gracilis to homogeneity as judged by polyacrylamide gel electrophoresis, using a novel citrate-Sepharose procedure. The enzyme was found to have a molecular weight of 520,000 and consisted of four identical subunits (Mr = 120,000). Two minor low molecular weight bands were observed on SDS-PAGE (Mr 51,000 and 49,000). Trypsin digestion experiments indicated that these could have been the result of limited proteolysis by an endogenous trypsin-like proteinase. In this respect, it resembles the mammalian ATP:citrate lyase. The enzyme was stimulated by NH+4 ions and inhibited by palmitoyl, lauroyl, oleoyl, myristoyl and stearoyl-CoA esters, glutamate and glucose 6-phosphate but not by acetyl-CoA or shorter chain fatty acyl-CoA esters. The enzyme exhibited normal Michaelis-Menten kinetics for citrate; however there was a 3-fold increase in Km with a high concentration of Cl- ions (0.25 M). The possible regulatory roles of ATP:citrate lyase in R. gracilis are discussed in the light of these findings.
Shashi
K
K
Department of Microbiology and Sanitation, Central Food Technological Research Institute, Mysore, India.
Bachhawat
A K
AK
Joseph
R
R
D001275:ATP Citrate (pro-S)-Lyase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D000214:Acyl Coenzyme A / Q000494:pharmacology; D000227:Adenine Nucleotides / Q000494:pharmacology; D002951:Citrates / Q000378:metabolism; D019343:Citric Acid; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D004952:Esters / Q000494:pharmacology; D007700:Kinetics; D046911:Macromolecular Substances; D003904:Mitosporic Fungi / Q000201:enzymology*; D008970:Molecular Weight; D011189:Potassium Chloride / Q000494:pharmacology; D012248:Rhodotorula / Q000201:enzymology*; D014357:Trypsin
D000214:Acyl Coenzyme A; D000227:Adenine Nucleotides; D002951:Citrates; D004952:Esters; D046911:Macromolecular Substances; D019343:Citric Acid; D011189:Potassium Chloride; D001275:ATP Citrate (pro-S)-Lyase; D014357:Trypsin